One of the mechanisms by which the body's immune system operates involves the engulfed of foreign protein by antigen presenting cells (APCs), where the proteins are broken down by proteases into peptide fragments, associated with the cell's MHC II protein in a peptide loading compartment, and then transported to the surface of the APC, where the peptide is presented in association with the MHC II to the T-cell receptors (TCRs) of T-helper cells (CD4.sup.+ T-cells). The TCRs of CD4 T-cells only recognise the antigenic peptides in association with MHC II, and the TCRs have a repertoire of recognition sites, so that only those helper cells with the appropriate TCRs will recognise a given antigenic peptide. This activates the helper cell to stimulate cytotoxic T-cells (CD8.sup.+ or Tc cells) and B-cells with the corresponding antigen specificity, which then mount an attack on the original source of foreign protein, either directly by CD4.sup.+ T-cells themselves, or by the production of antibody (B-cells) or by the Tc-cells. In the case of the latter, receptors on the Tc-cells recognise the original antigen presented in association with MHC I on the surface of cells in which they are endogenously produced, for example as a result of infection or through the generation of a mutated protein within the cell.
Vaccination has been of great importance in the protection against infectious diseases. Even so there is still a need to develop more safe and more effective vaccines. It is also necessary to develop vaccines for diseases for which there until now have been no such preventive measures. For effective vaccination, one needs to activate T-cells of the CD4 type because CD4.sup.+ T-cells direct activation of cytotoxic T-cells and B-cells.
Many cancer cells are the result of mutation, for example mutation in the p21 ras gene, and in consequence, an antigenic peptide fragment bearing the mutation is presented at the surface of the cell. The body's immune system may normally deal with such potential cancer cells by receptors on CD4.sup.+ T-cells recognising the apparently "foreign" antigen on MHC II and subsequently providing "signal 2" for specific cytotoxic T-cells which may kill the tumour cells. In this way, we probably fight off potential cancers by treating the cancer cells as though they had been infected with a foreign organism. However, in some cases tumours are established, and then it is beneficial to alter the balance between the T-cells and the tumour cells in favour of the T-cells by inducing or increasing specific T-cell activity.
The antigen peptide which locates in the MHC II molecule is typically about 11-20 amino acids in length, and one way of trying to efficiently vaccinate against infectious agents or to activate T-cells specific for tumour specific antigens, is to provide the peptide artificially. However, it has not proved very successful, probably because the peptide is easily degraded.
One group of workers (Zaghouani et al, Science (1993), 259:224-227, and Brumeanu et al, J Exp Med (1993), 178:1795-1799) disclosed the substitution of DNA encoding a viral epitope peptide into the DNA encoding the CDR3 loop of the heavy chain of an immunoglobulin molecule. This gene was co-expressed with a light chain to produce a complete Ig type of molecule, which was taken up by the Fc receptor (FcR) of an APC, and the viral epitope was presented with MHC II on the surface of the APC in vitro. The authors suggested that antigenized self Ig molecules could represent an effective carrier for delivery of peptides to MHC II molecules, as in vaccination or (tolerization) protocols, and a carrier that has a potential to be long-lived and devoid of side-effects.
We have studied antigen presentation of the 91-101 amino acid fragment from the .lambda.2 light chain of M315 antibody (Bogen et al, Eur J Immunol (1986), 16:1373). We have further used in vitro mutagenesis to move the epitope to loops in the human IgG3 heavy chain. Three different mutants were made, each with one of the loops of the CH1 domain replaced with the 91-101 peptide. The mutant genes were transfected into a fibroblast cell line which had previously been transfected with genes encoding the E.sub..alpha..sup.k E.sub..beta..sup.d MHC II. The resulting clones were assayed for the ability to stimulate .lambda.2.sup.315 specific T-cell clones. It appears that the mutated heavy chains are retained intracellularly in the transfected fibroblasts, but nevertheless the peptide is processed and presented on MHC II.